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A Genome-Scale DNA Repair RNAi Screen Identifies SPG48 as a Novel Gene Associated with Hereditary Spastic Paraplegia

Max Planck Institute for Molecular Cell Biology and Genetics, Dresden, Germany.
PLoS Biology (Impact Factor: 11.77). 06/2010; 8(6):e1000408. DOI: 10.1371/journal.pbio.1000408
Source: PubMed

ABSTRACT DNA repair is essential to maintain genome integrity, and genes with roles in DNA repair are frequently mutated in a variety of human diseases. Repair via homologous recombination typically restores the original DNA sequence without introducing mutations, and a number of genes that are required for homologous recombination DNA double-strand break repair (HR-DSBR) have been identified. However, a systematic analysis of this important DNA repair pathway in mammalian cells has not been reported. Here, we describe a genome-scale endoribonuclease-prepared short interfering RNA (esiRNA) screen for genes involved in DNA double strand break repair. We report 61 genes that influenced the frequency of HR-DSBR and characterize in detail one of the genes that decreased the frequency of HR-DSBR. We show that the gene KIAA0415 encodes a putative helicase that interacts with SPG11 and SPG15, two proteins mutated in hereditary spastic paraplegia (HSP). We identify mutations in HSP patients, discovering KIAA0415/SPG48 as a novel HSP-associated gene, and show that a KIAA0415/SPG48 mutant cell line is more sensitive to DNA damaging drugs. We present the first genome-scale survey of HR-DSBR in mammalian cells providing a dataset that should accelerate the discovery of novel genes with roles in DNA repair and associated medical conditions. The discovery that proteins forming a novel protein complex are required for efficient HR-DSBR and are mutated in patients suffering from HSP suggests a link between HSP and DNA repair.

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Available from: Mirko Theis, Aug 21, 2015
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    • "Disruption of spg11 (SPG11, spatacsin) or zfyve26 (SPG15, spastizin) expression during zebrafish development induced a range of developmental defects, including locomotor impairment and abnormal architecture of spinal MN axons (Martin et al., 2012; Southgate et al., 2010). These two proteins are components of a multi-protein complex (Słabicki et al., 2010) and AMOs targeting the two genes at the same time suggested that they were involved in a pathway required for spinal MN axon outgrowth (Martin et al., 2012). However, an EMS (Raldú a et al., 2008) was apparently induced by changing the expression of these genes, thus making it difficult to interpret the phenotype, which may also be due to a developmental delay. "
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    • "Consequently, the competition between homologous recombination and NHEJ must be considered when developing therapeutic applications based on the generation of specific DSBs using molecular scissors. Using RNAi, it was recently shown that the absence of proteins involved in non-conservative NHEJ DSB repair augments the frequency of repair events mediated by the conservative homologous recombination pathway in mammalian cells (Slabicki et al., 2010). This observation suggests that inhibition of NHEJ may favor DSB repair via the conservative mechanism. "
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    • "Several genomewide RNAi screening analyses have recently improved our understanding of the DNA repair pathway [52] [53] [54]. Słabicki et al. identified 61 genes affecting DNA DSB repair in human HeLa cells [54]. The downregulation of 17 of these genes led to an increase in endonuclease-induced homologous recombination. "
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